Project Summary/Abstract The corticospinal pathway is a pivotal mediator of voluntary motor control in humans and restoring corticospinal motor function after spinal cord injury remains one of the most substantial challenges facing translational and clinical neuroscience today. Regardless of the approach taken to alleviate the interruption of corticospinal axons, functional recovery will rely on the plasticity of the cortical motor network to incorporate the remodeled or replaced circuit. The creation of novel therapeutic interventions for the recovery of function after injury will have to be coupled to an understanding of the cellular and subcellular mechanisms that support cortical motor network remodeling and incorporation of injured neurons. Only recently have the tools necessary to answer these critical questions been developed. The long-term goal is to develop novel therapeutic interventions for the recovery of function after spinal cord injury through an understanding of the cellular and subcellular mechanisms that drive neural circuit remodeling. The overall objective for this proposal is to identify neuromodulatory mechanisms of motor map plasticity that correlate with recovery of skilled motor function and to determine the relationship between cortical network changes and corticospinal circuit remodeling after cervical spinal cord injury. The central hypothesis is that cholinergic input directly to corticospinal motor neurons is required for functional integration of corticospinal circuit changes into de novo motor networks after spinal cord injury. The rationale for the proposed research is that once the role of cholinergic activity in rehabilitation after spinal cord injury is defined, it is likely to provide new opportunities for pharmacological modulation and pairing with treatments that enhance corticospinal axon regeneration. The first approach will be to use cytotoxic lesions and optogenetic/chemogenetic control of relevant subcortical circuitry to modulate known mediators of motor learning while using two-photon microscopy to assess the functional incorporation of injured corticospinal neurons by measuring activity and structural changes during rehabilitation in awake, behaving animals. The second approach will be to define the molecular mechanisms and the cellular location of signaling events that underlie motor learning, and likely motor rehabilitation. The proposed studies are innovative in that they shift the focus of spinal cord rehabilitation onto the circuit mechanisms of cortical network plasticity and will have far- reaching importance in translating treatments for both acute and chronic injuries to motor networks. The proposed studies are significant because they will elucidate the mechanisms by which circuit remodeling influences recovery, which will have far-reaching importance in translating treatments for both acute and chronic injuries to cortical motor networks. The expectation is that completion of the proposed research will generate new opportunities for pharmacological modulation and combinatorial treatments that enhance corticospinal axon regeneration while mediating cortical motor network reorganization.